Use of the polymerase chain reaction and denaturing gradient gel electrophoresis to study diversity in natural virus communities

Abstract

Viruses are abundant members of marine and freshwater microbial communities, and are important players in aquatic ecology and geochemical cycles. Recent methodological developments have allowed the use of the polymerase chain reaction (PCR) to examine the diversity of natural communities of viruses without the need for culture. DNA polymerase genes are highly conserved and are, therefore, suitable targets for PCR analysis of microbes that do not encode rRNA. As natural virus communities are largely made up of dsDNA viruses, and as many dsDNA algal viruses encode their own DNA polymerase, PCR primers can be designed to amplify fragments of these genes. This approach has been used to examine the genetic diversity in natural communities of viruses that infect phytoplankton. Algal-virus-specific primers were used to amplify polymerase fragments from natural virus samples, demonstrating the presence of a diverse community of viruses closely related to those that are known to infect phytoplankton. We have modified this approach by using denaturing gradient gel electrophoresis (DGGE) to rapidly analyze PCR products. DGGE will permit rapid and efficient fingerprinting of natural marine viral communities, and allow spatial and temporal differences in viral community structure to be examined. This paper provides a brief overview of how PCR and DGGE can be used to examine diversity in natural viral communities drawing on viruses that infect phytoplankton as an example.